Our major goal is to develop a novel technology to identify HLA class II tumor antigens using a unique cloning approach in filamentous phages which has not been applied to the cloning of class II antigens in the past. This approach has numerous potential advantage over existing approaches to class II antigen cloning or biochemical peptide isolation. The unique cloning approach proposed here will be applicable to a variety of tumor-associated HLA class II antigens derived from tumors of different histologic types, and also to class II antigens involved in infectious diseases. Active specific immunotherapy has great potential for the treatment of patients with melanoma or colorectal carcinoma (CRC). In light of the important role that CD4+, HLA class II-dependent helper T (TH) cells play in the control of tumor growth, approaches to active immunotherapy with TH cell-defined antigens need to be developed. Very few tumor- associated TH antigens or peptides with vaccine potential have been identified thus far. The long-term goal of the proposed studies is to identify TH antigens with vaccine potential for melanoma and CRC patients. The Th antigens are recognized by available Th lines and clones. A novel approach for the cloning of mammalian HLA class II-dependent Th antigens will be developed using filamentous phages. Tumor cells cDNA libraries will be expressed by the phages, followd by library phage presentation to TH cell by antigen- presenting cells and identification of the relevant Th antigen in cytokine release assay. To develop this approach, we have available a unique model system including Th cells against tetanus toxoid and a cDNA fragment encoding the tetanus toxoid-associated Th epitope. Specifically we will: 1) develop Th antigen cloning technology using filamentous phages and tetanus toxoid as a model system (R21 pilot study); and 2) optimize the filamentous phage HLA class II antigen cloning approach developed under the pilot study to clone melanoma and CRC-associated Th antigens; this will be followed by the identification and characrterization of these antigens (R33). The feasibility of the filamentous phage approach for the cloning of HLA class II antigens is emphasized by the demonstration of successful induction of antigen-specific, Th cell-dependent antibodies in mice by immunizing the mice with phages expressing various antigens. The proposed studies will develop a novel technology to identify HLA class II-dependent Th antigens using filamentous phages leading to novel approaches to active specific cancer immunotherapy using these antigens. The antigens also may be of diagnostic value.